Ornithine decarboxylase (ODC), one of the most unstable protein, is ATP-dependently degraded by the 26S proteasome without ubiquitination. The degradation is dependent on antizyme, a polyamine-induced protein that binds to ODC,inhibiting its activity. The present study aimed to understand molecular mechanism whereby 26 S proteasome recognizes and degrades ODC-antizyme complex. First, we examined the role of antizyme by (1) spectrophotometric analysis of the structural changes of ODC caused by binding with antizyme, (2) analysis of functional regions of antizyme and analysis of degradation of mutant ODC.Second, we examined the site of ODC recognized by 26 S proteasome by (1) analysis of degradation products of ODC,(2) analysis of inhibitory effects of various synthetic peptides, each simulating different part of ODC sequence, on ODC degradation, and (3) identification of the enzyme which catalyses constitutive ODC degradation. The results suggested that antizyme binds to ODC monomer and inactivates it with C-terminal half (122-218) and elicits a conformational change of ODC with an adjacent region (113-118), resulting in exposure of a hidden degradation signal of ODC to 26 S proteasome. The multicatalytic proteinase was seemed to cleave ODC at many sites (mainly carboxyl sides of neutral/hydrophobic amino acid residues) by its endoproteolytic function, generating oligopeptides consisting of 5-11 amino acid residues. Antizyme was not degraded during ODC degradation. It was suggested that antizyme does not carry degradation signal within its molecule but accelerates constitutive ODC degradation, also catalyzed by the 26 S proteasome, by enhancing the association of ODC to proteinase. The degradation signal of ODC remained to be clarified by future work.